Preparation of beta-tricalcium phosphate microsphere-hyaluronic acid-based powder gel composite as a carrier for rhBMP-2 injection and evaluation using long bone segmental defect model

The specific objective of this study was to evaluate whether rhBMP-2-loaded bio-scaffolds can be used as effective rhBMP-2 carriers in the implantation of bone defect sites or poor bone quality in host bone. The rhBMP-2 release pattern test showed slow results in both groups, and a 1:9 ratio composition with a high water-absorption rate was selected for in vivo study. All animals euthanized after 9 weeks. The new bone formation and bone quantity and quality of fibular samples were examined. The results showed that the rhBMP-2 powder gel composite improved the new bone formation in the cortical bone and the marrow space. The length of new bone formation ratio of the rhBMP-2 loaded composite group was significantly higher than the powder gel group. The composite of powder gel seems to be a nice carrier, and slow release of rhBMP-2 can promote new bone formation in a segmental cortical bone defect after implantation.     

Enhanced bone regeneration at a segmental bone defect by controlled release of bone morphogenetic protein-2 from a biodegradable hydrogel

The objective of this study is to investigate the feasibility of a biodegradable hydrogel of gelatin as the controlled release carrier of bone morphogenetic protein-2 (BMP-2) suitable for enhancement of bone regeneration at a segmental bone defect. Hydrogels with three different water contents were prepared through glutaraldehyde crosslinking of gelatin with an isoelectric point of 9.0 under varied reaction conditions. Segmental critical-sized defects (20 mm) were created at the ulnar bone of skeletally mature New Zealand white rabbits, and gelatin hydrogels incorporating BMP-2 (17 microg/hydrogel) were implanted into the defects. When bone regeneration was evaluated by soft x-ray observation and bone mineral density (BMD) measurement, the gelatin hydrogels incorporating BMP- 2 exhibited significantly high osteoinduction activity compared with that of free BMP-2, although the activity depended on the water content of the hydrogels. Significantly higher BMD enhancement was observed in the gelatin hydrogel with a water content of 97.8 wt% than that with the lower or higher water content. We concluded that the biodegradable gelatin hydrogel is a promising controlled release carrier of BMP-2 for bone regeneration at the segmental bone defect.     

中空羟基磷灰石复合rhBMP-2修复骨缺损过程的再血管化研究

目的　探讨和观察中空羟基磷灰石复合rhBMP-2在骨缺损修复过程的再血管化。  方法　将48只成年的新西兰雄性大白兔制作成桡骨骨缺损模型,随机分3组,各组分别植入以下材料：中空HA/ rhBMP-2复合人工骨、单纯中空HA人工骨、单纯rhBMP-2。植入后于4、8、12、16周分别注射99mTc-MDP进行放射性核素骨显像并监测骨缺损修复过程中再血管化情况,同时进行大体、X线、组织学观察。  结果　术后各时间段,中空HA/ rhBMP-2复合人工骨组在X线及放射性核素聚集强度明显高于单纯中空HA人工骨组(P<0.05) ,表现为成骨代谢活跃及早期的再血管化能力。  结论　中空HA/ rhBMP-2复合人工骨具有良好的骨缺损修复能力,成骨活性持久,再血管化能力强,有望成为一种理想的骨缺损修复材料。     

An injectable scaffold: rhBMP-2-loaded poly(lactide-co-glycolide)/hydroxyapatite composite microspheres

Poly(lactide-co-glycolide)/hydroxyapatite(50/50) (PLGA/HA(50/50)) composite microspheres were fabricated and treated with a mixture of 0.25 M NaOH aqueous solution and ethanol (v/v = 1/1) at 37 °C. The properties of untreated and treated PLGA/HA(50/50) composite microspheres were determined and compared. The results showed that the surface roughness, HA content and hydrophilicity of the treated PLGA/HA(50/50) composite microspheres increased with treatment time. However, the treatment time should be kept within 2 h in order to maintain the shape of the PLGA/HA(50/50) microspheres. At the same time, a degradation study showed that both the untreated and treated microspheres degraded gradually with time, with the treated microspheres degrading faster in the first 4 weeks. The rhBMP-2-loaded PLGA/HA(50/50) composite microspheres were prepared by solution dipping treated PLGA/HA(50/50) composite microspheres. Mouse OCT-1 osteoblast-like cells were cultured on the untreated, treated and rhBMP-2-loaded PLGA/HA(50/50) composite microspheres and the cell affinity of the various microspheres was assessed and compared. It was found that the surface-treated PLGA/HA(50/50) composite microspheres clearly promoted osteoblast attachment, proliferation and alkaline phosphatase activity. It was considered that the hydrophilicity, osteoconductivity and surface roughness were increased by the increase in the HA component, which facilitated cell growth. Moreover, the rhBMP-2 loaded on the treated PLGA/HA(50/50) composite microspheres could be slowly released and further enhanced osteoblast differentiation. The good cell affinity and enhanced osteogenic potential of the rhBMP-2-loaded PLGA/HA composite microspheres indicate that they could be used as an injectable scaffold.     

长骨节段性缺损修复重建的理论及应用进展

背景：长骨节段性骨缺损修复重建方法众多,各有优缺点,限制了其广泛应用,仍是一个未完美解决的骨科难题。 目的：综述近年国内外长骨节段性缺损的修复重建进展研究。 方法：第一作者应用计算机检索1990年1月至2012年12月PubMed数据库、中国期刊全文数据库、维普数据库有关长骨节段性缺损修复重建的文章,英文检索词“bone defect, long bone reconstruction, tissue engineering of bone,scaffolds,bone reconstruction,bone graft,bone tumor; tumor resection,musculoskeletal tumors,regeneration,autografts”;中文检索词“大段,骨缺损,骨肿瘤,软组织肿瘤,瘤段切除,组织工程,骨移植”。共检索到104篇相关文献,纳入52篇文献进一步研究。 结果与结论：传统长骨节段性骨缺损的修复方法包括自体骨移植、异体骨移植、骨延长、人工假体置换等。研制适用于儿童保肢需要,符合儿童生长发育特点的假体成为儿童保肢的一个挑战课题。为了解决这一问题,人们研制出可以不断延长的假体,用于儿童骨肿瘤的保肢重建。随着医疗技术的进步,各种人工骨,尤其是骨组织工程和基因治疗在骨缺损修复中的逐步应用将进一步提高骨缺损的修复效果,是未来的研究方向。     

Alginate hydrogel linked with synthetic oligopeptide derived from BMP-2 allows ectopic osteoinduction in vivo

Bone morphogenetic proteins (BMP) are unique molecules with a specific biological activity for inducing ectopic bone formation when implanted with a suitable carrier matrix. However, incorporation of BMP into the carrier has disadvantages, including early burst release and protein degradation in biological environments. Therefore, we considered that the next greatest challenge in achieving successful clinical use was the development of a carrier system for site-specific delivery of the morphogenetic signal of BMP. In this study, a novel BMP-2-derived oligopeptide, NSVNSKIPKACCVPTELSAI, was coupled covalently to alginate. Then NSVNSKIPKACCVPTELSAI-linked alginate hydrogel composites were implanted into the calf muscle of rats and harvested 3 or 8 weeks after surgery. Ectopic bone formation was observed in alginate hydrogel linked with BMP-2-derived peptide. It is suggested that alginate hydrogel linked with an oligopeptide derived from BMP-2 might provide an alternative system for topical delivery of the morphogenetic signal of BMP-2.     

Ectopic osteoinduction and early degradation of recombinant human bone morphogenetic protein-2-loaded porous beta-tricalcium phosphate in mice

The present study investigated the ectopic osteoinduction and early degradation of recombinant human bone morphogenetic protein-2 (rhBMP-2)-loaded porous beta-tricalcium phosphate (beta-TCP) in mice. The porous beta-TCP with 50 microg of rhBMP-2 (n = 25) and porous beta-TCP (control group, n = 25) were implanted into muscle pouches in the right and left thigh of 28-day-old mice (n = 25), respectively. At every time point (3, 7, 14, 21 and 28 days after implantation), five mice were euthanized and the histological examinations of implantation sites were performed. In addition, the alkaline phosphatase (ALP) activity was also quantitatively analyzed. For the rhBMP-2-loaded group, blood vessel formation and immature cartilage was observed within the porous beta-TCP 3 days after implantation. Mature cartilage was observed 7 days after implantation of rhBMP-2-loaded porous beta-TCP. Newly formed woven bone, lamellar bone as well as marrow were observed 14 and 21 days after implantation of the rhBMP-2-loaded porous beta-TCP. Lamellar bone and marrow were observed 28 days after implantation of the rhBMP-2-loaded porous beta-TCP. For the control group, no bone or cartilage was observed at all time points. However, multinucleated giant cells and fibrous tissues were observed in the control group at 7 and 28 days after implantation, respectively. At 21 and 28 days after implantation, porous beta-TCP was observed to fragment indicating early degradation of the porous beta-TCP in both groups. In addition, ALP was observed to be significantly higher in the rhBMP-2-loaded beta-TCP as compared to the control beta-TCP. It was concluded from this study that the rhBMP-2-loaded porous beta-TCP induced blood vessel and ectopic bone formation.     

Injectable and thermo-sensitive PEG-PCL-PEG copolymer/collagen/n-HA hydrogel composite for guided bone regeneration

A novel three-component biomimetic hydrogel composite was successfully prepared in this study, which was composed of triblock PEG-PCL-PEG copolymer (PECE), collagen and nano-hydroxyapatite (n-HA). The microstructure and thermo-responsibility of the obtained PECE/Collagen/n-HA hydrogel composite were characterized. Scanning electronic microscopy (SEM) showed that the composite exhibited an interconnected porous structure. The rheological analysis revealed that the composite existed good thermo-sensitivity. In vivo biocompatibility and biodegradability was investigated by implanting the hydrogel composite in muscle pouches of rats for 3, 7, and 14 days. Moreover, the osteogenic capacity was evaluated by means of implanting the composite material in cranial defects of New Zealand White rabbits for 4, 12 and 20 weeks. In vivo performances confirmed that the biodegradable PECE/Collagen/n-HA hydrogel composite had good biocompatibility and better performance in guided bone regeneration than the self-healing process. Thus the thermal-response PECE/Collagen/n-HA hydrogel composite had the great potential in bone tissue engineering.     

钛网打压植骨重建四肢长骨节段骨缺损☆

背景：四肢长骨节段骨缺损临床上常见,一直以来是骨科修复领域的难题。 目的：介绍钛网打压植骨重建四肢长骨节段骨缺损新方法的实验研究和临床用用。 方法：第一作者应用计算机检索1990至2011年 PubMed数据库(http://www.ncbi.nlm.nih.gov/PubMed), CNKI 数据库(www.cnki.net/index.htm), 和维普数据库(http//www.cqvip.com)有关钛网打压植骨重建四肢长骨节段骨缺方面的文献,英文检索词为“segmental bone defect,titanium mesh or titanium cage”;中文检索词为“四肢或长骨,骨缺损,钛网”。排除重复性研究、非四肢长骨和Meta分析类文献。根据纳入标准,共检索到30篇文献进行归纳总结。 结果与结论：钛网打压植骨重建四肢长骨节段骨缺损无论在大动物或是小动物的实验中均取得较好的成骨效果,优于非钛网的游离植骨和大块皮质骨游离植骨;自体骨、异体骨和人工骨均是可行的填充植骨材料,以自体松质骨为主的植骨材料的成骨效果相对较好,具有生物活性钛网较普通钛网的成骨效果更好。临床上,在四肢长骨各个部位的节段骨缺损中均取得很好的治疗效果,包括骨缺损长达十余厘米的病例。钛网打压植骨重建四肢长骨节段骨缺损新方法,改变了以往认为大于 6 cm以上长段骨缺损只适宜带血运的骨移植、而不适宜无血运游离植骨的传统观点。该方法较简单、安全、有效,是一种较理想的替代治疗长骨骨干部节段骨缺损的无血运游离植骨法。     

Bone inductive properties of rhBMP-2 loaded porous calcium phosphate cement implants inserted at an ectopic site in rabbits

Recombinant human bone morphogenetic protein-2 (rhBMP-2) is known for its osteoinductive potential in bone tissue engineering. Calcium phosphate (Ca-P) cements are injectable, osteoconductive ceramic materials in which a macroporous structure can be induced during the setting reaction. In this study, the osteoinductive capability of rhBMP-2 loaded porous Ca-P cement was evaluated. Porous Ca-P cement discs were made and loaded with rhBMP-2 in vitro and implanted subcutaneously in the back of New Zealand white rabbits. The implantation period was either 2 or 10 weeks. Histological analysis of retrieved specimens revealed evident bone formation in the rhBMP-2 loaded Ca-P cement discs (pore fill: 18+/-6%) after 10 weeks of implantation. Bone formation occurred only in rhBMP-2 loaded porous Ca-P cement discs. Degradation of the Ca-P cement could not be confirmed after 10 weeks of implantation. The scaffold maintained its shape and stability during this time period. We conclude that porous Ca-P cement is a suitable carrier material for ectopic bone engineering.     

Biodegradable composite scaffolds incorporating an intramedullary rod and delivering bone morphogenetic protein-2 for stabilization and bone regeneration in segmental long bone defects

In this study, a two-part bone tissue engineering scaffold was investigated. The scaffold consists of a solid poly(propylene fumarate) (PPF) intramedullary rod for mechanical support surrounded by a porous PPF sleeve for osseointegration and delivery of poly(dl-lactic-co-glycolic acid) (PLGA) microspheres with adsorbed recombinant human bone morphogenetic protein-2 (rhBMP-2). Scaffolds were implanted into critical size rat segmental femoral defects with internal fixation for 12 weeks. Bone formation was assessed throughout the study via radiography, and following euthanasia, via microcomputed tomography and histology. Mechanical stabilization was evaluated further via torsional testing. Experimental implant groups included the PPF rod alone and the rod with a porous PPF sleeve containing PLGA microspheres with 0, 2 or 8 μg of rhBMP-2 adsorbed onto their surface. Results showed that presence of the scaffold increased mechanical stabilization of the defect, as evidenced by the increased torsional stiffness of the femurs by the presence of a rod compared to the empty defect. Although the presence of a rod decreased bone formation, the presence of a sleeve combined with a low or high dose of rhBMP-2 increased the torsional stiffness to 2.06 ± 0.63 and 1.68 ± 0.56 N·mm, respectively, from 0.56 ± 0.24 N·mm for the rod alone. The results indicate that, while scaffolds may provide structural support to regenerating tissues and increase their mechanical properties, the presence of scaffolds within defects may hinder overall bone formation if they interfere with cellular processes.     

Segmental bone regeneration using an rhBMP-2-loaded gelatin/nanohydroxyapatite/fibrin scaffold in a rabbit model

We aimed to develop a hybrid scaffold with a porous structure and similar composition as natural bone for the controlled release of bone morphogenetic protein-2 (BMP-2) to enhance bone regeneration. We fabricated a gelatin/nanohydroxypatite (nHAP) scaffold by glutaraldehyde chemical cross-linking a gelatin aqueous solution with nHAP granules at a 5:1 ratio (v/w). Then, fibrin glue (FG) mixed with recombinant human BMP-2 (rhBMP-2) was infused into the gelatin/nHAP scaffold and lyophilized to develop an rhBMP-2-loaded gelatin/nHAP/FG scaffold. On scanning electron microscopy, the composite had a 3-D porous structure. The rhBMP-2 release kinetics from the hybrid scaffold was sustained and slow, and release of rhBMP-2 was complete at 40 days. Immunohistochemistry, azo-coupling and alizarin S-red staining were used to study in vitro differentiation of human bone-marrow mesenchymal cells (hBMSCs). Strong positive staining results confirmed that rhBMP-2 released from the scaffold could improve osteocalcin (OCN) and alkaline phosphatase (ALP) expression and calcium deposition formation. RT-PCR results showed significantly high mRNA expression of ALP and OCN in hBM-MSCs cultured on the gelatin/nHAP/FG scaffold with rhBMP-2. DNA assay demonstrated that the scaffold was noncytotoxic and could promote hBMSC proliferation from the components of the hybrid scaffold, not released rhBMP-2. The hybrid scaffolds were then used to repair critical-size segmental bone defects of rabbit radius. Gross specimen, X-ray, bone histomorphology and bone mineral density assay demonstrated that the rhBMP-2-loaded gelatin/nHAP/FG scaffold had good osteogenic capability and could repair the segmental bone defect completely in 12 weeks.     

Repair of segmental long bone defect in rabbit femur using bioactive titanium cylindrical mesh cage

A segmental rabbit femur defect was repaired using an empty bioactive titanium (BAT) mesh cage. A 10mm long titanium mesh cage was positioned in the bony defect and reinforced by intramedullary fixation. The BAT surface was prepared by chemical and thermal treatment. Pure titanium cages were used as a control. Torsional stiffness of the BAT group at 4 weeks was approximately equal to, and at 8 weeks twice, that of the intact femur. Differences between the torsional stiffness of the control and BAT groups were significant at both time intervals. Histological examinations showed that woven bone appeared around the cage by 4 weeks and transformed to lamella bone by 8 weeks. New bone bonded to the BAT surface without an intervening layer. The BAT cage enhanced the bone repairing process and achieved faster repair of long bone segmental defects.     

Maxillary sinus floor elevation using a tissue-engineered bone with rhBMP-2-loaded porous calcium phosphate cement scaffold and bone marrow stromal cells in rabbits

The aim of this study was to evaluate the effects of maxillary sinus floor elevation by a tissue-engineered bone complex with recombinant human bone morphogenetic protein-2 (rhBMP-2)-loaded porous calcium phosphate cement (CPC) scaffold and bone marrow stromal cells (bMSCs) in rabbits. bMSCs were cultured and osteogenically induced. The osteoblastic differentiation of expanded bMSCs was detected by alkaline phosphatase activity, and calcium deposits in vitro. Thirty-six rabbits were randomly allocated into week 2, 4 and 8 observation groups. At each time point, 24 maxillary sinus floor elevation surgeries in 12 rabbits were performed bilaterally and randomly implanted by (1) CPC materials alone (group A, n = 6), (2) rhBMP-2/CPC composite materials alone (group B, n = 6), (3) CPC/bMSCs complex (group C, n = 6) and (4) rhBMP-2/CPC/bMSCs complex (group D, n = 6). As for maxillary sinus floor elevation, rhBMP-2-loaded CPC could promote new bone formation as compared to CPC, while addition of bMSCs could further enhance its new bone formation and maturity significantly, as detected by histological findings, and fluorochrome labeling. Our data suggested that rhBMP-2/CPC possessed excellent osteoinductive ability, while combining with bMSCs could further promote new bone formation and maturation in maxillary sinus elevation.     

Healing of segmental bone defects in rats induced by a beta-TCP-MCPM cement combined with rhBMP-2.

A beta-tricalcium phosphate-monocalcium phosphate monohydrate (beta-TCP-MCPM) cement was evaluated as an effective carrier of recombinant human bone morphogenetic protein-2 (rhBMP-2) in rat femoral critical-size defects. Hard cement cylinders (4 x 5 mm) impregnated with two different doses of rhBMP-2 (1.26 or 6.28 microg) were implanted into each defect, and the results were compared with those in rats that had implantations of cylinders only. Implantation of the 6.28 microg dose of rhBMP-2 caused a large bone shell to form around the defect, resulting in osseous union in all cases within 3 weeks. Except for beta-TCP granules, the cement was resorbed and replaced by bone tissue at 6 weeks. A torsion test at 9 weeks showed that the failure torque and bone stiffness had recovered 99% and 141%, respectively, compared with the intact contralateral femur. The defects that received 1.26 microg of rhBMP-2 resulted in 40% union and 41% of the failure torque at 9 weeks. However, no instances of union were observed in the defects implanted with cylinders only. In conclusion, the beta-TCP-MCPM cement was shown to be effective as a rhBMP-2 carrier. Combined with rhBMP-2, this cement was rapidly resorbed and completely healed the defects.     

A composite graft material containing bone particles and collagen in osteoinduction in mouse.

Demineralized allogenic bone matrices (DABM) and demineralized freeze-dried bone allograft (DFDBA) have been successfully used as bone-graft materials in the treatment of acquired and congenital cranio-maxillofacial defects and in some orthopedic surgery. However, these bone-graft "powders" have many shortcomings. For example, placement of particulate graft material in a hemorrhaging site can result in inadequacies or inaccurate attachment as well as loss of the graft materials. To minimize the inadequacies of powderlike graft materials, xenogenic collagen isolated from human tendon, skin, or bone was added to the bone-graft particles to form a composite spongelike implant. This material is commercially available and consists of 60% collagen and 40% DFDBA (DynaGraft, GenSci Co., Irvine, CA). The goal of this study was to evaluate the characteristics of composite graft implants in the mineralization process in an animal model in comparison with DFDBA powder and pure collagen. Seventy-two Swiss Webster mice were divided into three groups: an experimental group implanted with DynaGraft, two comparison groups implanted with either DFDBA or collagen only. All the graft materials were surgically implanted and inserted into the left thigh muscle. Mice were humanely killed at 1, 2, 3, 4, 6, 8, and 12 weeks. Then the muscle tissues in the vicinity of the implants were excised and processed for histology. Paraffin sections were stained with hematoxylin and eosin (H&E), the Von Kossa method, and Masson's trichrome. Some selected specimens were processed for transmission electron microscopic observation. After 1 week of implantation, the DynaGraft group showed calcium deposition on the collagen material and on the periphery of the DFDBA particles. Increased calcification and bone-forming cells were observed at 4-6 weeks. After 8 weeks, the implant formed a calcified nodule and only heavily mineralized connective tissue was observed at the implanted site. The group implanted with DFDBA powder showed calcification around the particulates. The collagen-sponge control group revealed no calcification or bone formation during the period of implantation. The light microscopic findings were confirmed by electron microscopy. Quantitative radiographic density DynaGraft and DFDBA graft followed sequentially over a period 120 days. It was concluded that a higher rate of calcification and bone formation was produced in the composite graft implant compared to the DFDBA implant. The composite graft material (DynaGraft), which contains both collagen and DFDBA, proved to be more effective for bone formation than particle components alone.     

Enhanced bone tissue regeneration by antibacterial and osteoinductive silica-HACC-zein composite scaffolds loaded with rhBMP-2

Next-generation orthopedic implants with both osteoinductivity and antibacterial ability are greatly needed. In the present study, biodegradable rhBMP-2 loaded zein-based scaffolds with a macroporous structure were synthesized, and SBA-15 nanoparticles and hydroxypropyltrimethyl ammonium chloride chitosan (HACC) were incorporated into the scaffolds to produce an anti-infective composite scaffold for delivery of osteogenic factors that facilitate the functional repair of bone defects. The silica/HACC/zein scaffolds developed here showed bioactivity, biocompatibility, and effective antibacterial activity. Confocal laser scanning microscopy (CLSM) was used to quantitatively measure the bactericidal efficacy with respect to bacterial adhesion. Results showed that the sample zein-HACC-S20 exhibited long-lasting antibacterial activity against Escherichia coli and Staphylococcus aureus up to 5 d. At a low dosage of rhBMP-2 (ca. 80 μg), the scaffolds released rhBMP-2 protein efficiently at a relatively slow rate, even after 27 d. An ALP activity and ECM mineralization assay showed that the zein-HACC-S20 scaffolds exhibited significant early osteogenic differentiation by generating enhanced ALP product on day 14 and ECM mineralization on day 21. In a mouse model of thigh muscle pouches, zein-S20 and zein-HACC-S20 groups resulted in obvious bone formation and gave more extensive mineralization to the implants than silica free groups, indicating effective bone induction in vivo. In a rabbit model of critical-sized radius bone defects (20 mm in length and 5 mm in diameter), the bone defects were almost fully repaired and bone marrow cavity recanalization was detectable by 3D micro-CT technique and histological analysis after 12 weeks. In this way, the zein-HACC-S20 scaffolds were proven to significantly promote the bone repair. They also demonstrated considerable promise for tissue engineering. Silica/HACC/zein scaffolds with both antibacterial activity and the ability to induce osteogenesis have immense potential in orthopedics and other biomedical applications.     

Improving bone formation in a rat femur segmental defect by controlling bone morphogenetic protein-2 release

Nonunion is a common complication in open fractures and other severe bone injuries. Recombinant human bone morphogenetic protein-2 (rhBMP-2) delivered on a collagen sponge enhances healing of fractures. However, the burst release of rhBMP-2 necessitates supra-physiological doses of rhBMP-2 to achieve a robust osteogenic effect, which introduces risk of ectopic bone formation and severe inflammation and increases the cost. Although the concept that the ideal pharmacokinetics for rhBMP-2 includes both a burst and sustained release is generally accepted, investigations into the effects of the release kinetics on new bone formation are limited. In the present study, biodegradable polyurethane (PUR) and PUR/microsphere [PUR/poly(lactic-co-glycolic acid)] composite scaffolds with varying rhBMP-2 release kinetics were compared to the collagen sponge delivery system in a critical-sized rat segmental defect model. Microcomputed tomography analysis indicated that a burst followed by a sustained release of rhBMP-2 from the PUR scaffolds regenerated 50% more new bone than the collagen sponge loaded with rhBMP-2, whereas a sustained release without the burst did not form significantly more bone than the scaffold without rhBMP-2. This study demonstrated that the putative optimal release profile (i.e., burst followed by sustained release) for rhBMP-2 can be achieved using PUR scaffolds, and that this enhanced pharmacokinetics regenerated more bone than the clinically available standard of care in a critical-sized defect in rat femora.     

Assessment of the initial viscoelastic properties of a critical segmental long bone defect reconstructed with impaction bone grafting and intramedullary nailing

IntroductionThis study compared the initial viscoelastic properties of a segmental tibial defect stabilized with intramedullary nailing and impaction bone grafting to that of a transverse fracture stabilized with intramedullary nailing.Materials and methodsSeven sheep tibiae were tested in compression (1000 N), bending and torsion (6 Nm) in a six degree-of-freedom hexapod robot. Tests were repeated across three groups: intact tibia (Intact), transverse fracture stabilized by intramedullary nailing (Fracture), and segmental defect stabilized with a nail and impaction bone grafting (Defect). Repeated measures ANOVA on the effect of group on stiffness/phase angle were conducted for each loading direction.ResultsThe Intact group was significantly stiffer than the Fracture and Defect groups in bending and torsion (p < 0.022 for both loading directions), and was marginal for the Defect group in compression (p = 0.052). No significant differences were found between the Fracture and Defect groups (p > 0.246 for all loading directions) for stiffness/phase angle. In compression and bending, phase angles were significantly greater for the Fracture and Defect groups compared to Intact (p < 0.025), with no significant differences between groups in torsion (p = 0.13). Sensitivity analyses conducted between the Fracture and Defect group differences found that they were not of clinical significance.ConclusionThe initial properties of a segmental defect stabilized with intramedullary nailing and impaction bone grafting was not clinically significantly different to that of a transverse fracture stabilized with intramedullary nailing.     

Reduction of ectopic bone growth in critically-sized rat mandible defects by delivery of rhBMP-2 from kerateine biomaterials

Absorbable collagen sponges (ACS) are used clinically as carriers of recombinant human bone morphogenetic protein 2 (rhBMP-2) to promote bone regeneration. ACS exhibit ectopic bone growth due to delivery of supraphysiological levels of rhBMP-2, which is particularly problematic in craniofacial bone injuries for both functional and esthetic reasons. We hypothesized that hydrogels from the reduced form of keratin proteins (kerateine) would serve as a suitable alternative to ACS carriers of rhBMP-2. The rationale for this hypothesis is that keratin biomaterials degrade slowly in vivo, have modifiable material properties, and have demonstrated capacity to deliver therapeutic agents. We investigated kerateine hydrogels and freeze-dried scaffolds as rhBMP-2 carriers in a critically-sized rat mandibular defect model. ACS, kerateine hydrogels, and kerateine scaffolds loaded with rhBMP-2 achieved bridging in animals by 8 weeks as indicated by micro-computed tomography. Kerateine scaffolds achieved statistically increased bone mineral density compared to ACS and kerateine hydrogels, with levels reaching those of native bone. Importantly, both kerateine hydrogels and kerateine scaffolds had significantly less ectopic bone growth than ACS sponges at both 8 and 16 weeks post-operatively. These studies demonstrate the suitability of keratins as rhBMP-2 carriers due to equal regenerative capacity with reduced ectopic growth compared to ACS.